Affinity Protein Purification - Day 1

One of the more efficient methods to enrich or purify a protein from other proteins and components in a crude cell lysate or other samples is affinity purification, whereby the protein of interest is purified by taking advantage of its specific binding properties to an immobilized ligand. It is important to select the optimal method for your purification project. To maintain a competitive advantage, many issues need to be addressed such as fusion tags, support for affinity purification, quality and efficiency. This conference addresses key questions such as: Which resins are optimal for which procedure? Do tags need to be removed and if so, how? What are the newest tagging technologies? Is there an optimal way to ensure high yields, high speed and low costs?

Affinity tags are widely used in protein science. Some are used to boost expression (both soluble and insoluble), some to increase solubility and many to specifically capture target proteins, especially when poorly expressed. In my talk I shall describe the tagging strategies we use for isolating recombinant proteases and related proteins, as well as the advantages and disadvantages of using such strategies.

1:45 Self-Cleaving Affinity Tags: A New Platform for Biologics Manufacturing?

The development of self-cleaving affinity tag technology has the potential to provide a new platform for biologics manufacturing, and several systems have been developed. The use of these technologies, however, will require an understanding of their impact on widely accepted downstream processes, as well as new validation methods for safe adoption and regulatory approval. This talk will examine these issues, with an emphasis on the specific opportunities for self-cleaving tags, as well as the remaining barriers to their application. This examination will include examples of how existing canonical processes might be impacted by the adoption of self-cleaving tag methods.

Sponsored by2:15 The three A’s of Biologicals are Activity, Activity, Activity: Case Study of Guiding Process Development Towards Biological Specific Activity though Bioactivity and in Vitro Kinetic AssaysOren Beske, Ph.D., Vice President, in Vitro Services, Aragen BiosciencesMonitoring quality and activity of a biological are essential during manufacturing development. This study describes significant bioactivity loss after adoption to an industrial CHO platform. Using the ForteBio Octet, an in vitro binding assay was rapidly developed, revealing a faster dissociation constant for the inactive form. Alternative clones and upstream processes were developed to increase specific activity. Both the bioactivity assay and the ForteBio kinetic assay were used to track product quality during process development.

A heme tag imparts intense color to a target protein for visual tracking during expression in E. coli and purification. Heme tags also can be utilized in affinity purification and protein quantification. Developments in heme tags will be discussed including achievement of tagging in the E. coli cytoplasm as well as periplasm.

TAP coupled with mass spectrometry constitutes a powerful tool for the characterization of protein complex associated with a given target protein. This talk reviews several alternative TAP tags with improved efficiency and/or flexibility. In particular, a newly developed SBP-His tandem tag will be discussed in more detail.

4:15 Moderated Small-Group Breakout Discussions

Discussion groups give participants an opportunity to network and discuss important topics with colleagues from around the world. We set aside this time for conference attendees to interact around a focused discussion topic, get to know one another, and develop contacts. Meeting delegates select what discussion they would like to join from the list of topics provided, and the talk begins. The discussions are engaging, and a great way to network, exchange information, and establish future collaborations.